Department of Ocean Sciences, Memorial University of Newfoundland, St. John's, NL A1C 5S7, Canada.
Department of Life Sciences and Health, Faculty of Health Sciences, OsloMet-Oslo Metropolitan University, N-0130 Oslo, Norway.
Cells. 2019 Dec 7;8(12):1592. doi: 10.3390/cells8121592.
MicroRNAs (miRNAs) are key regulators in fish immune responses. However, no study has previously characterized the impact of polyriboinosinic polyribocytidylic acid (pIC) and formalin-killed typical (ASAL) on miRNA expression in Atlantic salmon fed a commercial diet with and without immunostimulant CpG. To this end, first, we performed small RNA deep sequencing and qPCR analyses to identify and confirm pIC- and/or ASAL-responsive miRNAs in the head kidney of salmon fed a control diet. DESeq2 analyses identified 12 and 18 miRNAs differentially expressed in pIC and ASAL groups, respectively, compared to the controls. Fifteen of these miRNAs were studied by qPCR; nine remained significant by qPCR. Five miRNAs (miR-27d-1-2-5p, miR-29b-2-5p, miR-146a-5p, miR-146a-1-2-3p, miR-221-5p) were shown by qPCR to be significantly induced by both pIC and ASAL. Second, the effect of CpG-containing functional feed on miRNA expression was investigated by qPCR. In pre-injection samples, 6 of 15 miRNAs (e.g., miR-181a-5-3p, miR-462a-3p, miR-722-3p) had significantly lower expression in fish fed CpG diet than control diet. In contrast, several miRNAs (e.g., miR-146a-1-2-3p, miR-192a-5p, miR-194a-5p) in the PBS- and ASAL-injected groups had significantly higher expression in CpG-fed fish. Multivariate statistical analyses confirmed that the CpG diet had a greater impact on miRNA expression in ASAL-injected compared with pIC-injected fish. This study identified immune-relevant miRNA biomarkers that will be valuable in the development of diets to combat infectious diseases of salmon.
微小 RNA（miRNA）是鱼类免疫反应的关键调节因子。然而，以前没有研究描述聚肌胞苷酸（pIC）和福尔马林灭活的典型弧菌（ASAL）对大西洋鲑在商业饮食中添加免疫刺激剂 CpG 时 miRNA 表达的影响。为此，首先，我们进行了小 RNA 深度测序和 qPCR 分析，以鉴定和确认在食用对照饮食的鲑鱼头肾中 pIC 和/或 ASAL 反应性 miRNA。DESeq2 分析鉴定出与对照组相比，pIC 和 ASAL 组分别有 12 个和 18 个 miRNA 差异表达。通过 qPCR 研究了其中的 15 个 miRNA，其中 9 个通过 qPCR 仍然具有显著性。qPCR 显示 5 个 miRNA（miR-27d-1-2-5p、miR-29b-2-5p、miR-146a-5p、miR-146a-1-2-3p、miR-221-5p）被 pIC 和 ASAL 显著诱导。其次，通过 qPCR 研究了含 CpG 的功能性饲料对 miRNA 表达的影响。在预注射样本中，在喂食 CpG 饮食的鱼中，有 6 个 miRNA（例如 miR-181a-5-3p、miR-462a-3p、miR-722-3p）的表达明显低于对照组。相比之下，在 PBS 和 ASAL 注射组中，许多 miRNA（例如 miR-146a-1-2-3p、miR-192a-5p、miR-194a-5p）在喂食 CpG 的鱼中表达明显更高。多变量统计分析证实，CpG 饮食对 ASAL 注射鱼的 miRNA 表达的影响大于 pIC 注射鱼。本研究确定了与免疫相关的 miRNA 生物标志物，这将对开发防治鲑鱼传染病的饮食具有重要价值。
